Virtually all eukaryotic organisms appropriately examined have been shown to possess the capacity for endogenous temporal control and organization known as a circadian rhythm. The cellular machinery responsible for generating this ability is collectively known as the biological clock. The importance of a detailed understanding of the circadian clock to our understanding of physical and mental health and to the treatment of mental illness rests on the ubiquity of its influence on human mental and physiological processes. These range from circadian changes in basic physiological functions to the clear involvement of rhythms in human work rest cycles and sleep. Human psychiatric illnesses known to be a direct result of clock malfunction include forms of sleep phase disorders, manic-depressive illness and insomnia. Extensive research has demonstrated the significance of clock control of gene expression, but little is known regarding how clocks control the behavior of the cells in which they operate. One salient aspect of this regulation in all eukaryotes is the interplay of multiple feedback loops to control many aspects of cell physiology, including mRNA abundance. We have identified several such clock-interconnected loops that couple output to input and output back to the clock itself. We will use genetic and biochemical approaches to identify the cis- and trans-acting factors mediating this clock regulation. In Specific Aim 1 we will follow up the finding in Neurospora that a critical cell cycle regulator is clock-controlled and is essential for an output oscillator coupled to the clock loop. Additionally we will clone and characterize additional clock genes already identified in a genetic screen. In Specific Aim 2 we will examine the role of VVD in influencing output as well as input and PAS kinases in the clock, as well as carrying out a bioluminescence-based screen for factors affecting output. We have developed a strong conceptual and practical base in mammalian circadian biology and have begun the elucidation, via microarrays, of clock-regulated gene expression patterns in a mammalian cellular oscillator. In Specific Aim 3 we will exploit the identities of mammalian clock-regulated genes to identify regulatory pathways controlling circadian cell behavior. These approaches will be illuminated by the insights afforded through our work with both a simple eukaryotic model system and a mammalian cell culture model of the clock.